Department of Public Health, Chung Shan Medical University, Taichung 402, Taiwan; Division of Chest Medicine, Department of Internal Medicine, Changhua Christian Hospital, Changhua 500, Taiwan.
Oxidative Stress Group, Department of Environmental Health Sciences, and Biomolecular Sciences Institute, Florida International University, Miami, FL 33199, USA.
Redox Biol. 2019 Jan;20:556-565. doi: 10.1016/j.redox.2018.11.016. Epub 2018 Nov 26.
A reliable and fast liquid chromatography-tandem mass spectrometry method has been developed for the simultaneous determination of three oxidized nucleic acid damage products in urine, 8-oxoguanine (8-oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and 8-oxo-7,8-dihydroguanosine (8-oxoGuo). We applied this method to assess the effect of various urine workup procedures on the urinary concentrations of the oxidized nucleic acid products. Our results showed that frozen urine samples must be warmed (i.e., to 37 °C) to re-dissolve any precipitates prior to analysis. We showed that common workup procedures, such as thawing at room temperature or dilution with deionized water, are not capable of releasing fully the oxidized nucleic acid products from the precipitates, and result in significant underestimation (up to ~ 100% for 8-oxoGua, ~ 86% for both 8-oxodGuo and 8-oxoGuo). With this method, we further assessed and compared the ability of the three oxidized nucleic acid products, as well as malondialdehyde (MDA, a product of lipid peroxidation), to biomonitor oxidative stress in vivo. We measured a total of 315 urine samples from subjects with burdens of oxidative stress from low to high, including healthy subjects, patients with chronic obstructive pulmonary disease (COPD), and patients on mechanical ventilation (MV). The results showed that both the MV and COPD patients had significantly higher urinary levels of 8-oxoGua, 8-oxodGuo, and 8-oxoGuo (P < 0.001), but lower MDA levels, compared to healthy controls. Receiver operating characteristic curve analysis revealed that urinary 8-oxoGuo is the most sensitive biomarker for oxidative stress with area under the curve (AUC) of 0.91, followed by 8-oxodGuo (AUC: 0.80) and 8-oxoGua (AUC: 0.76). Interestingly, MDA with AUC of 0.34 failed to discriminate the patients from healthy controls. Emerging evidence suggests a potential clinical utility for the measurement of urinary 8-oxoGuo, and to a lesser extent 8-oxodGuo, which is strongly supported by our findings.
已开发出一种可靠且快速的液相色谱-串联质谱法，可同时测定尿液中三种氧化核酸损伤产物：8-氧鸟嘌呤（8-oxoGua）、8-氧-7,8-二氢-2'-脱氧鸟苷（8-oxodGuo）和 8-氧-7,8-二氢鸟苷（8-oxoGuo）。我们应用该方法评估了各种尿液处理程序对氧化核酸产物尿液浓度的影响。结果表明，冷冻尿液样本在分析前必须加热（即 37°C）以重新溶解任何沉淀物。结果表明，常见的处理程序，如室温解冻或用去离子水稀释，不能将氧化核酸产物从沉淀物中完全释放出来，从而导致显著的低估（8-oxoGua 高达约 100%，8-oxodGuo 和 8-oxoGuo 均约 86%）。使用该方法，我们进一步评估和比较了三种氧化核酸产物以及丙二醛（MDA，脂质过氧化产物）的能力，以监测体内氧化应激。我们共测量了 315 份来自低至高氧化应激负荷的受试者的尿液样本，包括健康受试者、慢性阻塞性肺疾病（COPD）患者和机械通气（MV）患者。结果表明，MV 和 COPD 患者的 8-oxoGua、8-oxodGuo 和 8-oxoGuo 尿液水平明显高于健康对照组（P<0.001），但 MDA 水平较低。受试者工作特征曲线分析显示，尿液 8-oxoGuo 是最敏感的氧化应激生物标志物，曲线下面积（AUC）为 0.91，其次是 8-oxodGuo（AUC：0.80）和 8-oxoGua（AUC：0.76）。有趣的是，AUC 为 0.34 的 MDA 未能区分患者和健康对照者。新出现的证据表明，测量尿液 8-oxoGuo 及其在较小程度上 8-oxodGuo 具有潜在的临床应用价值，这一发现得到了我们研究结果的有力支持。
